Currently, most analyte and DNA diagnostic assays are blood-based. Blood-based testing is invasive and poses a significant safety hazard. An alternate procedure, collecting lymph fluid, often involves surgically draining or removing lymph nodes. Collecting vaginal, rectal and nasal fluids is inconvenient and can be uncomfortable and embarrassing for both the medical professional and the patient. In contrast, the collection of oral fluid including saliva and/or mucosal transudate for testing entails minimal invasion of privacy, is convenient, relatively safe, and can be accomplished rapidly with ease. In addition, in patients where only small volumes of blood can be drawn, for example, newborns and the elderly, adequate volumes for testing can be obtained from saliva or mucosal transudate.
The idea for using oral fluid in a detection method has been discussed in scientific and clinical research for some time. Researchers have investigated using oral fluid as a possible clinical specimen for diagnosis of specific disease states or altered metabolic activity (see, e.g., Ann. New York Acad. Sci., Vol 694: Saliva as a Diagnostic Fluid, Malamud and Tabak, eds., N.Y. Acad. Sci. Pub. (1993)). Moreover, studies have shown that antibodies have been detected in saliva samples (Archibald et al. J Clin Micro. 24:873-875 (1986); Parry et al. Lancet 2:72-75 (1987)).
A number of enzyme immunoassay (EIA) kits are currently available for the screening of serum (blood) specimens for antibodies. An FDA approved EIA kit is also available specifically for screening oral fluid specimens (in particular oral fluid specimens collected using the OraSure.RTM. HIV-1 oral specimen collection device).
Detection of nucleic acid sequences in oral fluids by polymerase chain reaction (PCR) amplification and then visualization through agarose gel electrophoresis or blotting techniques has also been described (see, e.g., Irwin, et al, Nature Biotechnology 14:1146-8 (1996); Richards, et al., Human Molecular Genetics 2(2):159-63 (1993)). However, the literature describes using saliva or buccal cells from scraping or brushing the inside of the cheek.
The simultaneous collection of an oral fluid sample which is the basis of diagnostic and genomic assays has never been described. Simultaneous collection would be useful when screening assay results must be confirmed in another type of assay, i.e., an HIV test. Simultaneous collection would also be useful to diagnose two or more different pathological conditions by different assays. Instead of having to collect two samples of biological fluid; the investigator could collect one sample and run the necessary assays on that sample. Finally, simultaneous collection by a single device would be useful for the detection of genetic markers demonstrating a predisposition to a certain disease and the detection of the diagnostic markers indicating the presence of that disease.
In the present invention, a bibulous pad is used to collect oral fluids. This technique has proven to be better than the methods described in the prior art which involve collecting cells with buccal brushes and swabs (Richards, et al., supra) which can be painful and oral rinses with distilled water or saline (Irwin, et al., supra). Both of these methods produce a lower yield of nucleic acids and nucleic acids of lower quality than the methods and systems claimed below.
In the present invention, an absorbent pad is contacted with oral fluid. The fluid is expressed from the pad and assayed for the presence of diagnostic marker, including immunological markers and drug and drug metabolites. A surprisingly pure and stable preparation of genomic DNA is then extracted from the pad. The exceptional purity of the DNA is not fully understood but may be due to the preservative solution used to store the pad or the pad may preferentially bind DNA or cells which contain the genomic DNA.